1. Field of the Invention
This invention resides in the field of protein separations in biochemical research, and in particular the chemical species and mixtures used as carrier ampholytes in isoelectric focusing.
2. Description of the Prior Art
Of the two principal classes of-resolving media used in isoelectric focusing, carrier ampholytes offer certain advantages over immobilized pH gradients. One advantage is a particularly wide range of isoelectric point variation among individual species of an ampholyte mixture. Another is the versatility that arises from the ability of the ampholytes to function either in the liquid phase or on a solid or semi-solid support medium such as polyacrylamide, agarose, and Sephadex. This versatility permits separations to be performed on a large scale as well as a small scale, making it convenient both for preparative separations and for use in the research laboratory.
The pH range of any particular ampholyte mixture determines the type of separation that can be performed and the degree of resolution among solutes in the sample mixture. While the typical carrier ampholyte mixture of the prior art offers a pH range that extends well into the acid and base ranges, the typical upper end of the range is about 10.0. The effectiveness of ampholytes towards the outer reaches of the range is also limited by the tendency of ampholyte pH gradients to shift over time. This shift of the gradient often causes the gradient to flatten at each end, particularly at the upper end above pH 9. This limits the mixtures that can be separated effectively, and is particularly detrimental to two-dimensional electrophoresis in this range when isoelectric focusing in carrier ampholytes is used as the first dimension.